Multi-organ chip capable of being freely assembled and reused and multi-organ model construction method

Abstract

The invention relates to a blood flow connection type multi-organ chip capable of being freely assembled and reused, the blood flow connection type multi-organ chip comprises a chip cover body, organ culture chambers, a chip matrix and a pressure-sensitive film, the chip matrix comprises a plurality of groups of organ culture units which are mutually independent, each group of organ culture units comprises at least two culture flow channels and a plurality of organ culture holes, and the culture flow channels are communicated with the pressure-sensitive film. The organ culture holes are sequentially distributed at intervals along the culture flow channel, the bottom end of each organ culture hole is communicated with the culture flow channel, each organ culture hole is used for accommodating an organ culture chamber, the culture flow channel is located at the bottom of the chip substrate, one end of the culture flow channel is communicated with the storage pool, and the other end of the culture flow channel is communicated with the connecting port; the chip cover body is arranged at the top of the chip base body and completely covers the storage pools and the organ culture holes, the pressure-sensitive film covers the bottom of the chip base body to seal the culture flow channels, and in each group of organ culture units, a circulation path is formed between at least two culture flow channels through the storage pools. The biological function of the human body is better simulated, and disease modeling and treatment strategy research are carried out.

Description

Technical Field

[0001] The present invention relates to the field of biomedical engineering technology, and in particular to a method for constructing a high-efficiency blood flow-connected multi-organ chip and a multi-organ model that is both freely assembled and reusable. Background Art

[0002] Humans are complex biological systems. Organs within the body do not exist in isolation but rather within a highly integrated and dynamically interacting environment. Changes in one organ can affect other downstream organs. Organs communicate and interact through the bloodstream, transmitting various signals (soluble factors, exosomes, cells, etc.) to maintain normal physiological function. The construction of multi-organ models can more comprehensively simulate physiological and pathological processes in the human body and recreate a more realistic in vivo microenvironment. These models are used in reproductive medicine, physiological and pathological modeling, pharmacokinetic / pharmacodynamic analysis, and toxicological evaluation.

[0003] Animal models have problems such as high price, low throughput, species differences and ethics. Traditional two-dimensional cell culture lacks a three-dimensional environment for cell-cell and extracellular matrix (ECM) interactions, and also lacks organ-specific functions. Three-dimensional models cannot simulate the dynamic microenvironment in the body, and the diffusion of signal factors and nutrients is also limited. Therefore, these models cannot accurately simulate the development process of human diseases and predict the clinical efficacy and side effects of drugs. This has led to the current causes and pathological mechanisms of some diseases being unclear, treatment effects being lower than expected, the conversion rate of drugs to clinical applications being low, and the R&D process being time-consuming and costly.

[0004] Organ chips can reproduce the body's multicellular structure, tissue-tissue interface, physical and chemical microenvironment, and blood perfusion in vitro, and conduct intercellular communication, thereby constructing integrated functional units at the tissue and organ level. In the biological context of human tissue and organ function, real-time, high-definition microscopic imaging and in vitro detection of the biochemical and metabolic activity of living cells are also possible. Therefore, the organ chip platform enables researchers to easily control and change single or combined local cells, molecules, chemical and biophysical parameters, and analyze how they promote the development and progression of human diseases and responses to treatment. However, most of the current commercial multi-organ platforms cannot achieve the co-culture and communication and interaction of more than two organs, and there are problems such as complex operation and high cost. Standardization and high compatibility have not yet been achieved. Summary of the Invention

[0005] The main purpose of the present invention is to address the above-mentioned problems and provide a method for constructing a high-efficiency blood-flow-connected multi-organ chip and a multi-organ model that is both freely assembled and reusable, so as to better simulate the biological functions of the human body, carry out research on disease modeling and treatment strategies, improve the accuracy of drug development and toxicity assessment, and promote the development of personalized medicine.

[0006] To achieve the above objectives, the first aspect of the present invention provides a freely assembled and reusable blood flow-connected multi-organ chip, which mainly comprises a chip cover, an organ culture chamber, a chip substrate, and a pressure-sensitive membrane. The chip substrate comprises multiple groups of independent organ culture units, each group of the organ culture units comprises at least two culture channels and multiple organ culture wells, the multiple organ culture wells are spaced and sequentially distributed along the culture channels, the bottom end of each organ culture well is connected to the culture channel, and each organ culture well is used to accommodate the organ culture chamber. The culture channel is located at the bottom of the chip substrate, one end of the culture channel is connected to the retention tank, and the other end of the culture channel is connected to the connection port, which serves as a liquid inlet and a liquid outlet. The chip cover is arranged on the top of the chip substrate and completely covers the retention tank and the organ culture wells. The pressure-sensitive membrane covers the bottom of the chip substrate to seal the culture channel. In each group of organ culture units, a circulation path is formed between the at least two culture channels through the retention tank.

[0007] Preferably, all the culture channels of each group of the organ culture units share a storage tank, and the organ culture holes extend upward from the bottom to the upper surface of the chip substrate.

[0008] Preferably, the chip base has a first part and a second part, the height of the first part is greater than the height of the second part, the connection ports are evenly distributed in the second part, the retention tank and organ culture well are located in the first part, and the chip cover is snapped onto the first part.

[0009] Preferably, the number of organ culture wells is 16, 36 or 64, and the arrangement and size correspond to commercial cell culture 24-well plates, 48-well plates, and 96-well plates, and are compatible with detection equipment and match cell culture devices; the number of culture flow channels is 4, 6 or 8, the number of retention tanks is 1, 2, 3 or 4, and the number of connection ports is 4, 6 or 8.

[0010] Preferably, the width of each of the culture flow channels is the same as the diameter of the organ culture well disposed therein.

[0011] Preferably, the storage tank and the connection port are both connected to the culture flow channel via a triangular-shaped buffer structure.

[0012] Preferably, the width of the retention tank is 5 mm, and the connection port has a threaded design for connecting to a peristaltic pump or a syringe pump.

[0013] Preferably, the organ culture chamber has a cylindrical chamber body, the bottom surface of the cylindrical chamber body is a porous membrane, and a sealing ring is arranged around the circumference of the cylindrical chamber body. The sealing ring makes the upper and lower regions of the multilayer membrane independent of each other, and substances are exchanged only through the multilayer membrane. The top of the organ culture chamber is symmetrically provided with grooves for inserting and removing the organ culture chamber and assembling it with a tissue scaffold.

[0014] The tissue scaffold is one or two of a gel scaffold, a microplate scaffold, a porous scaffold and an elastic scaffold;

[0015] The pore size of the porous membrane is 0.4 μm to 8 μm.

[0016] Preferably, the thickness of the pressure-sensitive film is 10 μm to 300 μm.

[0017] A second aspect of the present invention provides a method for constructing a multi-organ model based on the blood flow connected multi-organ chip, the main feature of which is that the method comprises the following steps:

[0018] (1) Building a vascular barrier

[0019] A preparation containing basement membrane components was pre-laid on the lower layer of the porous membrane of the organ culture chamber. After gelation at 37°C, cells were plated at a density of 0.1 to 1 × 10 6 The endothelial cell suspension of 100 cells / mL was inoculated into the lower layer of the porous membrane and placed in an incubator for culture. After the cells adhered to the wall, the vascular barrier was constructed.

[0020] (2) Constructing organ substance

[0021] Select organ-forming cells and tissue scaffolds, and perform two-dimensional or three-dimensional cell culture of the selected organ on the porous membrane of the organ culture chamber to complete the construction of the organ's substantial part;

[0022] (3) Construction of multi-organ models

[0023] Based on the volume ratio and circulation sequence of the selected organs, the number and arrangement order of each organ module are determined, and the organ culture chambers corresponding to each organ are assembled on the chip substrate and connected to the fluid device for perfusion culture to complete the construction of the multi-organ model.

[0024] Compared with the prior art, the present invention has the following beneficial technical effects:

[0025] 1) The operation is simple. The overall size of the chip and the size and position of the organ culture wells are consistent with those of commercial well plates, making it compatible with a variety of automated culture or detection equipment.

[0026] 2) By adjusting the number and position of each organ culture chamber, the actual volume ratio and circulation process of each organ in the body are simulated to better reproduce the in vivo microenvironment.

[0027] 3) The plug-and-play design of the organ culture chamber allows organ models to be constructed under specific culture conditions, and the connection between organs is completed through the chip's blood circulation channels in the form of organ modules.

[0028] 4) A storage pool is designed on the chip substrate to reduce the overall volume of the device and facilitate culture medium replacement and sampling in subsequent experiments.

[0029] 5) The connection port on the chip substrate adopts a threaded design to reduce the possibility of leakage. It is also compatible with a variety of fluid devices and can apply unidirectional, bidirectional or cyclic fluid shear force to the model.

[0030] 6) The application of pressure-sensitive film reduces the thickness of the bottom of the chip and improves light transmittance, making it easier to observe under a microscope and to subsequently dismantle, clean and disinfect it.

[0031] 7) The chip material should be biocompatible and high-temperature resistant materials to maintain long-term cell culture and make the organ chip reusable.

[0032] 8) The application of tissue scaffolds can better simulate the biochemical, mechanical and structural properties of the extracellular matrix, reproduce the organ-specific microenvironment, structure and function, and can not only meet the culture conditions of traditional two-dimensional and three-dimensional cell models, but also construct organoid models in organ culture chambers.

[0033] 9) Two or more organ models can be constructed and co-cultured simultaneously. BRIEF DESCRIPTION OF THE DRAWINGS

[0034] Figure 1 This is a schematic diagram of the structure of the freely assembled and reusable blood flow-connected multi-organ chip of the present invention.

[0035] Figure 2 This is a schematic diagram of the disassembly of the freely assembled and reused blood flow-connected multi-organ chip of the present invention.

[0036] Figure 3 This is a perspective view of the freely assembleable and reusable blood flow-connected multi-organ chip after assembly.

[0037] Figure 4 This is a schematic top view of the chip substrate in the freely assembleable and reusable blood flow-connected multi-organ chip of the present invention.

[0038] Figure 5 This is a schematic diagram of the structure of the organ culture chamber in the freely assembled and reused blood flow-connected multi-organ chip of the present invention.

[0039] Figures 6 to 9 Schematic diagram of the structure of various tissue scaffolds in the freely assembled and reused blood flow connected multi-organ chip of the present invention.

[0040] Figure 10 These are live-dead staining images of various organ models constructed in the freely assembleable and reusable blood flow-connected multi-organ chip of the present invention. DETAILED DESCRIPTION

[0041] In order to more clearly understand the technical content of the present invention, the following embodiments are given in detail. However, it should be noted that these descriptions are only for further illustrating the features and advantages of the present invention, and are not intended to limit the claims of the invention.

[0042] Unless otherwise specified, the reagents and methods involved in the examples are commonly used in the art.

[0043] like Figures 1 to 10 The figure shows a specific embodiment of the freely assembleable and reusable blood-flow-connected multi-organ chip of the present invention. It facilitates culture and testing, and also offers the advantages of reusability, high compatibility, and simple operation. It is suitable for research in reproductive medicine, physiology, and pathology modeling, pharmacokinetic / pharmacodynamic analysis, and toxicology evaluation.

[0044] The multi-organ chip comprises a chip cover 1, an organ culture chamber 2, a chip base 3, and a pressure-sensitive film 4. The chip cover 1 is positioned on top of the chip base 3, while the pressure-sensitive film 4 covers the bottom of the chip base 3. The pressure-sensitive film 4 has a thickness of 10 μm to 300 μm and can be sealed and easily removed from the chip base, facilitating microscopic observation and post-use disinfection.

[0045] In this embodiment, the chip substrate 3 includes two groups of independent organ culture units, each group of the organ culture units includes two culture channels 32 and eight organ culture wells 31, and every four organ culture wells 31 are spaced and distributed in sequence along the corresponding culture channels 32. The bottom end of each organ culture well 31 is connected to the culture channel 32. Each organ culture well 31 is used to accommodate an organ culture chamber 2. The culture channel 32 is located at the bottom of the chip substrate 3. A storage tank 33 is provided at one end of the culture channel 32, and a connecting port 34 is provided at the other end of the culture channel 32. The connecting port 34 serves as a liquid inlet and also a liquid outlet. Figure 3 As shown, the chip cover 1 completely covers the reservoir 33 and organ culture well 31. A pressure-sensitive membrane covers the bottom of the chip substrate to seal the culture flow channel. In each group of organ culture units, a circulation path is formed between the two culture flow channels through the reservoir.

[0046] like Figure 1 As shown, all two culture channels 32 of each group of organ culture units share a reservoir 33, which reduces the overall volume of the device and facilitates culture medium replacement and sampling in subsequent experiments. The organ culture well 31 extends upward from its bottom end to the upper surface of the chip substrate 3.

[0047] like Figure 2 and Figure 3 As shown, the chip base 3 has a first part and a second part. The height of the first part is greater than that of the second part. The connection ports 34 are evenly distributed in the second part. The storage tank 33 and the organ culture hole 31 are located in the first part. The chip cover 1 is buckled on the first part to ensure that each organ is cultured under sterile conditions.

[0048] In addition to the above embodiments, the chip substrate 3 can also include 3 or 4 groups of independent organ culture units, each group of organ culture units includes 2 or more culture channels 32, and each culture channel 32 can also be provided with 6 or 8 organ culture wells 31. That is, the number of organ culture wells can be 16, 36, or 64, and the arrangement and size correspond to commercial cell culture 24-well plates, 48-well plates, and 96-well plates. It is compatible with various detection equipment such as microplate readers and microscopes, and matches various automated pipetting cell culture devices. The number of organ culture units, culture channels, and organ culture wells can be adjusted as needed.

[0049] like Figure 4 As shown, the width of each culture channel 32 is the same as the diameter of the organ culture well 31 disposed therein, and the positions correspond to each other. The width of the reservoir 33 can be 5 mm, with the width of the reservoir 33 parallel to the length of the chip substrate. The connection port 34 has a threaded design for connection to a peristaltic pump or syringe pump, reducing the possibility of leakage. It is also compatible with a variety of fluid devices and can apply unidirectional, bidirectional, or cyclic fluid shear forces to the model. Alternatively, it can be placed directly on a gravity shaker to provide a stable and continuous fluid environment for organ culture.

[0050] like Figure 1 As shown, the storage tank 33 and the connection port 34 are connected to the culture channel 32 via a triangular buffer structure 35. That is, both ends of the culture channel 32 have a buffer structure to connect to the storage tank and the connection port.

[0051] like Figure 5As shown, the organ culture chamber 2 has a cylindrical chamber body 21, the bottom surface of the cylindrical chamber body 21 is a porous membrane, and a sealing ring 22 is arranged around the circumference of the cylindrical chamber body. The pore size of the porous membrane is 0.4μm to 8μm. The porous membrane can allow the transport of cell metabolites and drugs, and realize the interactive connection between various organs. Multiple sealing rings 22 are evenly spaced and distributed. The top of the organ culture chamber 2 is symmetrically provided with grooves 23 for inserting and removing the organ culture chamber and assembling it with the tissue scaffold 5. The organ culture chamber of the present invention is different from commercialized In comparison, the culture channel at the bottom of the chip can form an independent closed space, and the area of ​​the bottom porous membrane is larger, which is conducive to the mutual communication between organs. It can ensure that the upper and lower layers of the porous membrane are cultured independently, but not closed. The diameter of the central porous membrane is smaller than the diameter of the culture hole in the orifice plate. In the present invention, the sealing ring is used to make the upper and lower regions of the multilayer membrane independent of each other, and only through the multilayer membrane to exchange substances. Specifically, the culture chamber designed by the present invention makes the upper and lower layers completely independent with the help of the sealing ring. They are in a closed state. The multilayer membrane does not circulate up and down. Only the lower region is located in the culture flow channel. The upper region is connected to the culture flow channel only through the multilayer membrane, that is, substances can only be exchanged with the help of the holes on the multilayer membrane. The upper layer is used to culture the organ substance, and the lower layer is used to construct the vascular barrier. This avoids the upper and lower liquids that may affect each other during the operation and also increases the bottom area. The various parts interact only through the porous membrane, and the organ substance of the upper layer is separated from the vascular barrier of the lower layer.

[0052] The organ culture chambers 2 are installed in the organ culture wells, and the actual volume ratio and circulation process of each organ in the body can be simulated by adjusting the number and position of different organ culture chambers 2 .

[0053] Figure 6 It is shown that the elastic scaffold is suitable for the culture of organs with mechanical stress, such as the heart; Figure 7 The gel scaffold is shown to be suitable for the culture of organs with barrier functions such as the lungs and intestines, or organs with three-dimensional structures such as the brain; Figure 8 The microplate scaffold is shown to be suitable for culturing organs with three-dimensional structures such as the liver; Figure 9 Porous scaffolds are suitable for culturing organs such as bone that require a three-dimensional microenvironment. Tissue scaffolds can act as an extracellular matrix to regulate cell behavior, function, and properties.

[0054] Each organ culture chamber 2 contains only one specific organ, namely, the various cells and specific tissue scaffolds that make up the organ. Depending on the model structure and functional characteristics, the tissue scaffold can be one or a combination of gel scaffolds, microplate scaffolds, porous scaffolds and elastic scaffolds to construct a corresponding two-dimensional or three-dimensional cell model.

[0055] The preparation materials of the chip cover 1, the main body of the organ culture chamber 2 and the chip substrate 3 can be one of the high-temperature resistant materials with good biocompatibility such as PC, PMMA, COC or COP. The porous membrane material of the organ culture chamber 2 can be one of PDMS, PC or PET. The material of the pressure-sensitive membrane 4 can be a highly permeable and high-temperature resistant material such as PET. The tissue scaffold 5 can be a natural material, a synthetic material or a mixed material, which is mainly selected according to the extracellular matrix composition and structural and functional characteristics of the organ.

[0056] The present invention provides a method for constructing a multi-organ model based on the aforementioned blood-flow-connected multi-organ chip. The constructed organ model comprises a parenchymal portion and a vascular portion. The organ can be a combination of physiological or pathological models of two or more organs, such as the intestine, liver, kidney, heart, lung, brain, pancreas, and bone, to construct a multi-organ model. This model is suitable for research in reproductive medicine, physiological and pathological modeling, pharmacokinetic / pharmacodynamic analysis, and toxicological evaluation. Each organ model is constructed in a separate organ culture chamber and then assembled with the chip substrate. The connection between organs is achieved through the chip's blood circulation channels.

[0057] The method specifically comprises the following steps:

[0058] (1) Building a vascular barrier

[0059] A preparation containing basement membrane components was pre-laid on the lower layer of the porous membrane in the organ culture chamber 2. After gelation at 37°C, cells were plated at a density of 0.1 to 1×10 6 An endothelial cell suspension of 100 cell / mL is inoculated into the lower layer of the porous membrane and cultured in an incubator. After the cells adhere to the wall, the vascular barrier is constructed. Preparations containing basement membrane components include: fibrin, laminin, type I and type IV collagen, etc. After 6-8 hours of cell adhesion, the organ culture chamber is flipped over to complete the construction of the vascular barrier.

[0060] (2) Constructing organ substance

[0061] According to the functional and structural characteristics of each organ and the research purpose, organ-constituting cells and tissue scaffolds 5 can be selected, and two-dimensional or three-dimensional cell culture of the selected organ can be carried out on the upper layer of the porous membrane of the organ culture chamber 2 to complete the construction of the organ substance; the gel scaffold can be used as a two-dimensional coating or a three-dimensional scaffold, and the microporous plate scaffold, porous scaffold and elastic scaffold can be used alone or in combination with the gel scaffold for three-dimensional cell culture.

[0062] (3) Construction of multi-organ models

[0063] According to the volume ratio and circulation sequence of each selected organ, the number and arrangement order of each organ module are determined, and the organ culture chamber 2 corresponding to each organ is assembled on the chip substrate 3 and connected to the fluid device for perfusion culture to complete the construction of the multi-organ model.

[0064] Example 1 In vitro serial connection of multi-organ physiological models and construction of pathological microenvironment

[0065] The porous membrane of the organ culture chamber is pre-laid with a preparation containing basement membrane components, including fibrin, laminin, type I and type IV collagen, etc. After gelation at 37°C, cells with a density of 0.1 to 1×10 6 An endothelial cell suspension of 100 cells / mL is inoculated onto the lower layer of the porous membrane and cultured in an incubator. After 6-8 hours of cell attachment, the organ culture chamber is flipped over to complete the construction of the vascular barrier. Based on the functional and structural characteristics of each organ and the research objectives, organ-constituting cells and tissue scaffolds are selected. Two-dimensional or three-dimensional cell culture of the organ is performed on the upper layer of the porous membrane of the organ culture chamber to complete the construction of the organ's parenchyma. For example, the intestinal model uses intestinal epithelial cells and a gel scaffold for two-dimensional culture, the liver model uses liver and Kupffer cells and a gel scaffold or a gel scaffold and a microplate scaffold for three-dimensional culture, the lung model uses lung epithelial cells and a gel scaffold for two-dimensional culture, the kidney model uses renal tubular epithelial cells and a gel scaffold for two-dimensional culture, the brain model uses neurons, astrocytes, and pericytes and a gel scaffold for three-dimensional culture, and the bone model uses osteoblasts and osteoclasts and a gel scaffold or a porous scaffold for three-dimensional culture. Then, according to the volume ratio and circulation sequence of each organ in the body, the number and arrangement order of each organ module are determined, and each organ culture chamber is assembled with the chip and connected to the fluid device for perfusion culture to complete the construction of the multi-organ series physiological model. Cell viability of the organ model is evaluated using cell viability staining reagents, such as Figure 10 Finally, disease models are constructed by adding disease-related pathogenic factors or virulence factors; for example, inflammation-pathogenic bacteria or metabolites, diabetes-high sugar environment, and hepatic steatosis-free fatty acids.

[0066] Example 2 Drug Evaluation

[0067] Taking the efficacy and toxicity evaluation of breast cancer drugs as an example, a multi-organ model of intestine-liver-heart-breast cancer series was constructed.

[0068] Fibrin was pre-laid on the lower layer of the porous membrane of the organ culture chamber. After gelation at 37°C, cells were cultured at a density of 0.1 to 1 × 10 6A suspension of human umbilical vein endothelial cells (HUVECs) at 100 cells / mL was inoculated onto the lower layer of the porous membrane and placed in an incubator for culture. After 6-8 hours of cell attachment, the organ culture chamber was flipped over to complete the construction of the vascular barrier. Intestinal, liver, heart, and breast cancer models were constructed on the upper layer of the porous membrane in the organ culture chamber. The intestinal model used intestinal epithelial cells and a gel scaffold for two-dimensional culture, the liver model used liver and Kupffer cells, and a gel scaffold or a gel scaffold and a microplate scaffold for three-dimensional culture, the heart model used cardiomyocytes, and a gel scaffold or a gel scaffold and an elastic scaffold for two-dimensional or three-dimensional culture, and the breast cancer model used breast cancer cells and a gel scaffold for three-dimensional culture. Based on the volume ratio of each organ in the body and the circulation order of the intestine, liver, heart, and breast tumors, the number and arrangement order of each organ module were determined. Each organ culture chamber was assembled with a chip and connected to a fluidic device for perfusion culture to complete the construction of a multi-organ serial breast cancer model. Finally, breast cancer drugs (e.g., doxorubicin) were added to the upper layer of the intestinal model to evaluate and analyze the drug's absorption in the intestine, metabolism in the liver, efficacy, and toxicity to each organ.

[0069] In this specification, the present invention has been described with reference to specific embodiments thereof. However, it will be apparent that various modifications and variations may be made without departing from the spirit and scope of the present invention. Therefore, the description is to be regarded as illustrative rather than restrictive.

Claims

1. A freely assembleable and reusable blood flow connected multi-organ chip, characterized in that: The chip substrate comprises a plurality of independent organ culture units, each of which comprises at least two culture channels and a plurality of organ culture holes. The plurality of organ culture holes are spaced and arranged in sequence along the culture channel. The bottom end of each organ culture hole is connected to the culture channel. Each organ culture hole is used to accommodate the organ culture chamber. The culture channel is located at the bottom of the chip substrate. One end of the culture channel is connected to the storage tank, and the other end of the culture channel is connected to the connecting port. The connecting port serves as a liquid inlet and a liquid outlet at the same time. The chip cover is arranged on the top of the chip substrate and completely covers the storage tank and the organ culture holes. The pressure-sensitive film covers the bottom of the chip substrate to close the culture channel. In each group of organ culture units, a circulation path is formed between the at least two culture channels through the storage tank.

2. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: All the culture channels of each group of organ culture units share a storage tank, and the organ culture holes extend upward from the bottom to the upper surface of the chip substrate.

3. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: The chip base has a first part and a second part, the height of the first part is greater than the height of the second part, the connection ports are evenly distributed in the second part, the retention tank and organ culture well are located in the first part, and the chip cover is buckled on the first part.

4. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: The number of organ culture wells is 16, 36, or 64, and the arrangement and size correspond to commercial cell culture 24-well plates, 48-well plates, and 96-well plates, and are compatible with detection equipment and match cell culture devices; the number of culture flow channels is 4, 6, or 8, the number of retention tanks is 1, 2, 3, or 4, and the number of connection ports is 4, 6, or 8.

5. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: The width of each of the culture flow channels is the same as the diameter of the organ culture well arranged therein.

6. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: The storage tank and the connection port are both connected to the culture flow channel through a triangular buffer structure.

7. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: The width of the storage tank is 5 mm, and the connection port has a threaded design for connecting to a peristaltic pump or a syringe pump.

8. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: The organ culture chamber comprises a cylindrical chamber body, the bottom surface of which is a porous membrane. A sealing ring is provided around the circumference of the cylindrical chamber body. The sealing ring makes the upper and lower regions of the multilayer membrane independent of each other, and substances are exchanged only through the multilayer membrane. Grooves are symmetrically provided on the top of the organ culture chamber for inserting and removing the organ culture chamber and assembling it with a tissue scaffold. The tissue scaffold is one or two of a gel scaffold, a microplate scaffold, a porous scaffold and an elastic scaffold; The pore size of the porous membrane is 0.4 μm to 8 μm.

9. The freely assembleable and reusable blood flow connected multi-organ chip according to claim 1, characterized in that: The thickness of the pressure-sensitive film is 10 μm to 300 μm.

10. A method for constructing a multi-organ model based on the blood flow connected multi-organ chip according to any one of claims 1 to 9, characterized in that: The method comprises the following steps: (1) Building a vascular barrier A preparation containing basement membrane components was pre-laid on the lower layer of the porous membrane of the organ culture chamber. After gelation at 37°C, cells were plated at a density of 0.1 to 1 × 10 6 The endothelial cell suspension of 100 cells / mL was inoculated into the lower layer of the porous membrane and placed in an incubator for culture. After the cells adhered to the wall, the vascular barrier was constructed. (2) Constructing organ substance Select organ-forming cells and tissue scaffolds, and perform two-dimensional or three-dimensional cell culture of the selected organ on the porous membrane of the organ culture chamber to complete the construction of the organ's substantial part; (3) Construction of multi-organ models Based on the volume ratio and circulation sequence of the selected organs, the number and arrangement order of each organ module are determined, and the organ culture chambers corresponding to each organ are assembled on the chip substrate and connected to the fluid device for perfusion culture to complete the construction of the multi-organ model.

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